Cancer is a major health challenge, with over 150,000 cases of cancer expected to be diagnosed in Canada in 2013 alone. While patients with early stage disease can be treated effectively by conventional therapies (surgery, radiation, chemotherapy), few options are available to patients with advanced disease and those options are typically palliative in nature. Active immunotherapy seeks to employ the patient's immune system to clear tumor deposits and offers an exciting option to patients who have failed conventional therapies (Humphries, 2013). Indeed, several clinical studies have demonstrated that immunotherapy with T cells can be curative in patients with advanced melanoma, confirming the utility of this approach (Humphries, 2013). Additionally, patients suffering from chronic lymphocytic leukemia (CLL) and acute lymphoblastic leukemia (ALL) have also been effectively treated and cured with T cell immunotherapy (Fry and Mackall, 2013) (Kochenderfer and Rosenberg, 2013). While there are several immunotherapy approaches, the engineering of T cells with chimeric receptors allows any patient's immune cells to be targeted against any desirable target in a major histocompatibility complex (MHC) independent manner. To date, the chimeric receptors used for engineering T cells consist of a targeting domain, usually a single-chain fragment variable (scFv); a transmembrane domain; and a cytosolic domain that contains signaling elements from the T cell receptor and associated proteins (Dotti et al., 2009). Such chimeric receptors have been referred to as “T-body”, “Chimeric Antigen Receptor” (CAR) or “Chimeric Immune Receptor” (CIR)—currently, most researchers use the term “CAR” (Dotti et al., 2009). These CARs are considered in modular terms and scientists have spent considerable time investigating the influence of different cytoplasmic signaling domains on CAR function. The first-generation CARs employed a single signaling domain from either CD3□ or Fc□RI□. Second-generations CARs combined the signaling domain of CD3□ with the cytoplasmic domain of costimulatory receptors from either the CD28 or TNFR family of receptors (Dotti et al., 2009). Third-generation CARs combined multiple costimulatory domains, but there is concern that third-generation CARs may lose antigen-specificity (Han et al., 2013). Most CAR-engineered T cells that are being tested in the clinic employ second-generation CARs where CD3□ is coupled to the cytoplasmic domain of either CD28 or CD137 (Han et al., 2013) (Finney et al., 2004) (Milone et al., 2009).
While CAR-engineered T cells have shown considerable promise in clinical application, they rely on a synthetic method for replacing the activation signal that is provided by the T cell receptor (TCR). Since this synthetic receptor does not deliver all of the signaling components associated with the TCR (ex. CD3epsilon, Lck), it remains unclear whether the T cells are optimally activated by the CAR or how the CAR activation affects T cell differentiation (ex. progression to memory). Furthermore, since the CAR signaling domains are disconnected from their natural regulatory partners by the very nature of the CAR structure, there is also an inherent risk that CARs may lead to a low-level of constitutive activation which could result in off-target toxicities.
Given these limitations, it is preferable to re-direct T cells to attack tumors via their natural TCR. To this end, a class of recombinant proteins termed “Bispecific T-cell Engagers” (BiTEs) has been created (Chames and Baty, 2009) (Portell et al., 2013). These proteins employ bispecific antibody fragments to crosslink T-cell TCR receptors with target antigens. This leads to efficient T-cell activation, triggering cytotoxicity. Similarly, bi-specific antibodies have been generated that accomplish this goal and some scientists have simply linked anti-CD3 antibodies to tumor-specific antibodies employing chemical linkage (Chames and Baty, 2009). While these bi-specific proteins have demonstrated some activity in vitro, GMP production, short biological half-lives and bioavailability represent significant challenges to the successful use of these molecules in cancer treatment. Additionally, these molecules also fail to properly recapitulate natural TCR signaling because they do not engage the TCR co-receptors (CD8 and CD4).
Accordingly, a need remains for T cell-antigen couplers with enhanced activity and safety compared to traditional CARs.